Pyrenoid loss in Chlamydomonas reinhardtii causes limitations in CO2 supply, but not thylakoid operating efficiency

The pyrenoid of the unicellular green alga Chlamydomonas reinhardtii is a microcompartment situated in the centre of the cup-shaped chloroplast, containing up to 90% of cellular Rubisco. Traversed by a network of dense, knotted thylakoid tubules, the pyrenoid has been proposed to influence thylakoid biogenesis and ultrastructure. Mutants that are unable to assemble a pyrenoid matrix, due to expressing a vascular plant version of the Rubisco small subunit, exhibit severe growth and photosynthetic defects and have an ineffective carbon-concentrating mechanism (CCM). The present study set out to determine the cause of photosynthetic limitation in these pyrenoid-less lines. We tested whether electron transport and light use were compromised as a direct structural consequence of pyrenoid loss or as a metabolic effect downstream of lower CCM activity and resulting CO₂ limitation. Thylakoid organization was unchanged in the mutants, including the retention of intrapyrenoid-type thylakoid tubules, and photosynthetic limitations associated with the absence of the pyrenoid were rescued by exposing cells to elevated CO₂ levels. These results demonstrate that Rubisco aggregation in the pyrenoid functions as an essential element for CO₂ delivery as part of the CCM, and does not play other roles in maintenance of photosynthetic membrane energetics.We wish to gratefully acknowledge financial support to ODC by Wolfson College, the Cambridge Philosophical Society and the TH Middleton Fund (Department of Plant Sciences) toward research-related travel, which was crucial to enable this collaborative study. This work was supported by the Biotechnology and Biological Sciences Research Council (PhD studentship 1090746 to ODC and BB/M007693/1 to MTM and HG). The work was also supported by NSF grant MCB 0951094 and US Department of Energy Grants DE-FG02-07ER64427, DE- FG02-12ER16338 awarded to ARG